OBJECTIVE: To determine whether nifedipine inhibits capacitative calcium entry at clinically relevant concentrations using cultured human myocytes as a model for human myometrium. METHODS: Myocyte cultures were initiated from the myometrium of term pregnant women who underwent cesarean delivery. Paired cells were chosen for study. The cell of interest was stimulated by an intercellular calcium wave from the adjacent cell. In this fashion, release of sarcoplasmic reticulum (SR) calcium was accomplished with minimal disturbance of the plasma membrane and the subplasmalemmal space (SPS) of the cell studied. Depletion of the SR calcium stores by the calcium wave activated the capacitative calcium current, elevated calcium in the SPS, and activated calcium-activated potassium channels. A cell-attached patch clamp was used to monitor the outward current resulting from the calcium activation of these potassium channels. Calcium green-1 fluorescence was used to simultaneously monitor changes of the deep cytosolic calcium concentrations. Experiments were performed at varying concentrations of nifedipine (0-10 micromol/L). RESULTS: Nifedipine reduced outward potassium currents in a dose-dependent manner. Nifedipine at 100 nmol/L resulted in greater than a 50% reduction of outward current, indicating a significant inhibition of capacitative calcium entry at that concentration. Higher concentrations of nifedipine abolished outward current. Experiments designed to detect indirect effects of nifedipine on capacitative calcium entry were negative. CONCLUSIONS: Nifedipine block of capacitative calcium entry occurred at concentrations similar to those required to block L-type voltage-activated calcium channels. These data suggest that block of capacitative calcium entry may be an important mechanism of action when nifedipine is clinically used for tocolysis of preterm labor.

Calcium↗

PubMedDate not supplied

Maintaining squamous epithelial architecture in a human cervical epithelium to SCID mouse xenograft model system.

OBJECTIVE: To establish a mouse model system of human cervical epithelium that maintains the stratified squamous architecture overlying a well-defined basement membrane. METHODS: Normal cervical tissue specimens were obtained from the transformation zone of hysterectomy specimens. Small (2 mm) cubes of tissue containing stroma and epithelium were placed subdermally into severe combined immunodeficient (SCID) mice and harvested between 7 and 45 days. Specimens were examined histologically for the presence of stroma, epithelium, and basement membrane. RESULTS: In 49 xenograft attempts in 24 mice, 35 (71%) were identified microscopically. Of these, 28 (80%) had identifiable epithelium and 20 (57%) formed cysts lined with stratified squamous epithelium. A cuboidal basal layer clearly defined the basement membrane. Proliferation within the cyst epithelium was demonstrated by expression of the Ki67 proliferation marker and the presence of mitotic figures. CONCLUSIONS: A human to mouse xenograft model system is described that maintains, in high yield, stratified squamous epithelium overlying a well-defined basement membrane. Advantages of this model system include the following: (1) The tissue originates from readily available human specimens; (2) A stratified squamous epithelial architecture is maintained for a sufficient time to study viral effects on proliferation; (3) The basement membrane of the epithelium is well defined, allowing for investigation of connective tissue factors in cervical intraepithelial neoplasia and cervical cancer.

Animals↗

PubMedDate not supplied

Isomers of a marine diterpene distinguish sublines of human melanoma cells on the basis of apoptosis, cell cycle arrest and differentiation markers.

The action of the marine furanoditerpenes, spongiatriol (SP) and episopongiatriol (ESP), were compared in two sublines of human melanoma cells (MM96E and MM96L) derived from the same metastatic lesion. MM96E had higher tyrosinase activity and lower expression of alkaline phosphatase but was otherwise indistinguishable from MM96L. SP and ESP treatment of both cell lines for 72 h at cytostatic doses inhibited B8G3 expression and tyrosinase activity but had little effect on the expression of tyrosinase antigen. MM96L cells were affected more than MM96E. SP and ESP induced apoptosis in both cell lines, ESP causing dendritic morphology in a proportion of MM96L cells. SP induced a marked G2/M arrest in MM96E cells. SP and ESP together define subtle qualitative and quantitative differences in human melanoma phenoypes, possibly based on expression of a repertoire of neurotransmitter receptors.

Antigens, Neoplasm↗

PubMedDate not supplied

Fentanyl and alfentanil initiate the migrating myoelectric complex but not the vomiting response in the canine small intestine.

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Action Potentials↗